The delivery of biologically active agents (including therapeutically relevant compounds) to subjects is often hindered by difficulties in the compounds reaching the target cell or tissue. In particular, the trafficking of many biologically active agents into living cells is highly restricted by the complex membrane systems of the cells. These restrictions can result in the need to use much higher concentrations of biologically active agents than is desirable to achieve a result, which increases the risk of toxic effects and side effects. One solution to this problem is to utilise specific carrier molecules which are allowed selective entry into the cell. Lipid carriers, biodegradable polymers and various conjugate systems can be used to improve delivery of biologically active agents to cells.
One class of biologically active agents that is particularly difficult to deliver to cells is a biotherapeutic (including nucleosides, nucleotides, polynucleotides, nucleic acids and derivatives). In general, nucleic acids are stable for only a limited duration in cells or plasma. The development of RNA interference, RNAi therapy, RNA drugs, antisense therapy and gene therapy, among others, has increased the need for effective means of introducing active nucleic acid agents into cells. For these reasons, compositions that can stabilise and deliver nucleic acid-based agents into cells are of particular interest.
The most well-studied approaches for improving the transport of foreign nucleic acids into cells involve the use of viral vectors or cationic lipids. Viral vectors can be used to transfer genes efficiently into some cell types, but they generally cannot be used to introduce chemically synthesized molecules into cells.
An alternative approach is to use delivery compositions incorporating cationic lipids which interact with a biologically active agent at one part and interact with a membrane system at another part (for a review, see Feigner, 1990, Advanced Drug Delivery Reviews, 5, 162-187 and Felgner, 1993, J. Liposome Res., 3, 3-16). Such compositions are reported to contain liposomes.
Since the first description of liposomes in 1965 by Bangham (J. Mol. Biol. 13, 238-252), there has been a sustained interest and effort in developing lipid-based carrier systems for the delivery of biologically active agents. The process of introducing functional nucleic acids into cultured cells by using positively charged liposomes is first described by Philip Felgner et al. Proc. Natl. Acad. Sci., USA, 84, 7413-7417 (1987). The process was later demonstrated in vivo by K. L. Brigham et al., Am. J. Med. Sci., 298, 278-281 (1989).
Liposomes are attractive carriers since they protect biological molecules from degradation while improving their cellular uptake. Out of the various classes of liposome, liposomes which contain cationic lipids are commonly used for delivering polyanions (e.g. nucleic acids). Such liposomes can be formed using cationic lipids alone and optionally including other lipids and amphiphiles such as phosphatidylethanolamine. It is well known in the art that both the composition of the lipid formulation as well as its method of preparation affect the structure and size of the resultant aggregate.
The use of cationic lipids for cellular delivery of biologically active agents has several advantages. The encapsulation of anionic compounds using cationic lipids is essentially quantitative due to electrostatic interaction. In addition, it is believed that the cationic lipids interact with the negatively charged cell membranes initiating cellular membrane transport (Akhtar et al., 1992, Trends Cell Bio., 2, 139; Xu et al., 1996, Biochemistry 35, 5616).
Following Felgner's early work on introducing functional nucleic acids into cultured cells by using positively charged liposomes, cationic lipid compounds based on general formula I have been disclosed in patent application EP 0 187 702.

These cationic lipid compounds generally consist of two alkyl or alkenyl chains linked to the nitrogen containing “head” group. There is also the disclosure of two or three of R3, R4 and R5 taken together being quinuclidino, piperidino, pyrrolidino or morpholino.
Since EP 0 187 702, various other researchers have disclosed cationic lipid compounds. A relevant patent application is WO00/030444 which describes synthetic cationic lipids and liposomes. The application discloses cationic lipid compounds with a variety of different head groups; some examples feature more than one head group. Amongst the compounds disclosed are compounds of formula (II).

WO 2005/121348 discloses lipid-based formulations. The nucleic acid-lipid particles disclosed therein comprise an interference RNA molecule, a cationic lipid with alkyl side chains from about 10 to 20 carbon atoms having more than a single site of unsaturation, a noncationic lipid and a conjugated lipid that inhibits aggregation of the particle such as a polyethyleneglycol (PEG)-lipid conjugate or a polyamide (ATTA)-conjugate. Specific cationic lipid compounds disclosed in this patent application include DSDMA, DODMA, DLinDMA, DLenDMA.

Cullis and Bailey, Biochemistry 1994, 33, 12573-12580, reports liposomes which comprise amino lipids such as the following.

Another recent publication concerning new cationic lipid compounds is WO2008/137758 which describes a range of amino acid lipid compounds reported as being useful for drug delivery and diagnosis.
US 2006/0240554 and related applications US 2008/0020058 and US 2009/0048197 also relate to cationic lipids. These lipids are reported to be capable of delivering biologically active agents, including small nucleic acid molecules such as short interfering nucleic acids (siNA) to cells and/or tissues.
US2006/0240554, US2008/0020058 and US2009/0048197 describe the use of the cationic lipid CLinDMA, in combination with cholesterol and PEG-DAG for delivering siRNA to cells. The structure of CLinDMA is set out below; at the “head” of the molecule is the —N(Me)2 group. ClinDMA is also referenced herein as E0173.

WO2009/086558 discloses the following compounds:

It also discloses amino lipids having the following general formula (in which R3 and R4 may join to form an optionally substituted heterocyclic ring of 4 to 6 carbon atoms and 1 or 2 heteroatoms chosen from nitrogen and oxygen):

There is a need for further cationic lipids which facilitiate the systemic and local delivery of biologically active agents to cells. There is also a need for cationic lipids which, relative to those cationic lipids that are known in the art, improve the systemic and local delivery of biologically active agents to cells. There is a further need for lipid formulations that have optimized physical characteristics for improved systemic and local delivery of biologically active agents to specific organs and to tumors, especially tumors outside the liver.